Pain from intra-articular NGF or joint injury in the rat requires contributions from peptidergic joint afferents

Abstract

Non-physiological stretch of the cervical facet joint's capsular ligament induces persistent behavioral hypersensitivity and spinal neuronal hyperexcitability via an intra-articular NGF-dependent mechanism. Although that ligament is innervated by nociceptors, it is unknown if a subpopulation is exclusively responsible for the behavioral and spinal neuronal responses to intra-articular NGF and/or facet joint injury. This study ablated joint afferents using the neurotoxin saporin targeted to neurons involved in either peptidergic ([Sar(9),Met (O2)(11)]-substance P-saporin (SSP-Sap)) or non-peptidergic (isolectin B4-saporin (IB4-Sap)) signaling to investigate the contributions of those neuronal populations to facet-mediated pain. SSP-Sap, but not IB4-Sap, injected into the bilateral C6/C7 facet joints 14 days prior to an intra- articular NGF injection prevents NGF-induced mechanical and thermal hypersensitivity in the forepaws. Similarly, only SSP- Sap prevents the increase in mechanical forepaw stimulation- induced firing of spinal neurons after intra-articular NGF. In addition, intra-articular SSP-Sap prevents both behavioral hypersensitivity and upregulation of NGF in the dorsal root ganglion after a facet joint distraction that normally induces pain. These findings collectively suggest that disruption of peptidergic signaling within the joint may be a potential treatment for facet pain, as well as other painful joint conditions associated with elevated NGF, such as osteoarthritis.

Keywords: Afferent; Facet joint; NGF; Neuronal hyperexcitability; Pain; Trauma.

Figure 1

Intra-articular NGF-induced behavioral hypersensitivity and neuronal hyperexcitability require joint afferents involved in peptidergic signaling. (A) Mechanical withdrawal threshold at day 1 is reduced from baseline for Blank-Sap+NGF (*p<0.004; n=12) and compared to SSP-Sap+ NGF (+p<0.001; n=12), SSP-Sap+veh (#p<0.001; n=8), and Blank-Sap+veh (‡p<0.001; n=4). (B) Thermal withdrawal latency is reduced for Blank-Sap+NGF at day 1 compared to baseline (*p<0.009; n=4) and SSP-Sap+NGF (+p<0.022; n=8) or SSP-Sap+veh (#p<0.011; n=8) but not Blank-Sap+veh (n=4). (C) Spinal neuronal extracellular recordings show greater evoked firing for Blank-Sap+NGF (n=4) than for SSP-Sap+NGF (+p<0.008; n=4) and SSP-Sap+ veh (#p<0.044; n=4) for 10g and 26g filaments (D). The proportion of wide dynamic range neurons (E) is not different between groups.

Figure 2

Ablating non-peptidergic joint afferents does not prevent intra-articular NGF-induced behavioral sensitivity or neuronal hyperexcitability. (A) Mechanical withdrawal threshold at day 1 is reduced compared to baseline for IB4-Sap+NGF (*p<0.001; n=8) and Saporin+NGF (#p<0.001; n=9); both exhibit lower thresholds than IB4-Sap+veh at day 1 (‡p<0.006; n=6). (B) Thermal withdrawal latency is reduced for IB4-Sap+NGF (*p<0.001; n=8) and Saporin+NGF (#p<0.001; n=9) compared to baseline, but only Saporin+NGF is lower than IB4-Sap+veh at day 1 (‡p<0.046; n=6). Spinal neuronal recordings (C) demonstrate the increased evoked firing (26g stimulation) for IB4-Sap+NGF (+p<0.038; n=5) and Saporin+NGF (^p<0.020; n=4) compared to IB4-Sap+veh (n=6) (D). The ratio of wide dynamic range neurons is not different between groups (E).

Figure 3

FJD-induced pain and increased NGF in the DRG utilizes joint afferents involved in peptidergic signaling. (A) Forepaw mechanical withdrawal threshold is reduced by day 1 and remains reduced through day 7 (p<0.001) after injury for Blank-Sap+FJD (n=7) and is lower than that for SSP-Sap+FJD (*p<0.001; n=7) or SSP-Sap+sham (#p<0.001; n=4) at all days. NGF expression and the percentage of neurons expressing NGF (NGF⁺) are each increased at day 7 in the DRG after Blank-Sap+FJD (n=7) (B) compared to SSP-Sap+FJD (*p<0.001; n=7) (C) and SSP-Sap+ sham (#p<0.001; n=4) (D) (E–F). 50µm scale bar in (D) applies to (B–D).